As a result, cells must acquire secondary mutations to allow proliferation and tumorigenesis in order to survive with BRCA1 deficiency (Aly and Ganesan, 2011). Almost all BRCA1-deficient cancer cells have acquired p53 mutations, but p53 function is not enough to overcome the growth defect resulted by BRCA1 loss. Losses of p53 can only delays embryonic lethality in full null BRCA1-mutant mice by a few days. Except this aspect, there are still several defects (Aly and Ganesan, 2011). These secondary mutations might interfere with the chemotherapeutic drug, the poly(ADP-ribose) polymerase (PARP) inhibitor, which lead to a great impact in cancer chemotherapy

Cao et al. first found this functional interaction between 53BP1 and BRCA1 by investigating the effect of mutations in other DNA repair and checkpoint proteins on BRCA111/11 mutant cells. 53BP1, a nuclear protein, is the human ortholog of yeast DNA damage checkpoint proteins Rad9p/Crb2, with a key role in DNA repair response and checkpoint control (Cao et al, 2009). Upon the induction of DNA DSBs, 53BP1 rapidly redistributes from a diffuse nuclear localization to discrete foci that co-localize with phosphorylated histone H2AX and other repair proteins including BRCA1. Cells lacking 53BP1 are sensitive to DNA damaging agents and have defects in both S-phase and G2M checkpoints (Mochan et al., 2003). Cao et al found that loss of 53BP1 could completely rescue the BRCA1-deficiency cells from mid-gestational embryonic lethality and tumorigenesis, by which increases the level of HR DSB repair.

With further experiment on ES cells with null-allele of Brca1, it extended the founding that loss of 53BP1 could rescue null-mutation of BRCA1 (Cao et al., 2009). Also they observed that loss of 53BP1 alleviated the spontaneous DNA damage, chromosomal abnormalities, and G2/M checkpoint activation associated with loss of BRCA1. Loss of 53BP1, but not p53, reversed the sensitivity of BRCA1-/- cells to cisplatin and mitomycin C. Moreover loss of 53BP1 restored Rad51 foci formation after IR treatment on BRCA1-/- cells, and partly restored HR function in BRCA1-/- as measured by gene targeting (Cao et al., 2009). There are three functions of 53BP1 that we already known; that is, transducing a subset of ATM-dependent cell-cycle checkpoints, facilitating the joining of distal DSBs formed at dysfunction telomeres, and during lymphocyte antigen receptor recombination (Bouwman et al. 2010). However, theses functions of 53BP1 cannot fully explain the mechanism of its reversed Brca1-deficient effect that was illustrated by Cao et al.

In this paper, Bunting et al. focus on the discussion that the BRCA1-deficient cells couldn’t perform the error-free HR on DSB because of the presence of 53BP1, and investigated the possible mechanism of the 53BP1 inhibitory function on HR (Bunting et al., 2010). Their studies showed that 53BP1 could block the resection of DNA ends at DSBs, which is a primary step of HR process. In order to provide evidences for this hypothesis, Bunting et al. firstly illustrated the observed phenomenon of loss of 53BP1, and obtained the evidence that loss of 53BP1 help to relieve the replication stress of aberration in S phase, which is induced by either PARPi or CPT, a topoisomerase I poisons (Bunting et al., 2010).

Then, they stated that this effect of loss of 53BP1 is due to increased HR activity in BRCA1 mutant cells by measuring three HR-related factors–Rad51 foci formation, sister chromatid exchanges, and DR-GFPhyg reporter system. When they had a closer look of the process of HR, they found out that 53BP1 only affect the activity of BRCA1, an early stage factor of HR, but not XRCC2, which is a downstream factor required for Rad51 foci formation in HR (Bunting et al., 2010). Additionally, they measured the RPA phosphorylation level, which dependent on ATM and CtlP activity at DSB resection, and found out that loss of 53BP1 shows a higher level (Bunting et al., 2010). Therefore, these experiment results support the hypothesis that 53BP1 affect the resection of DSB end in the primary step of HR.

Based on what is known, Bunting et al. proposed a possible mechanism for 53BP1 inhibitory function in HR. That is, in WT, 53BP1 binding to the DSB end to inhibit CtlP, assisted by BRCA1, to disassociate 53BP1 and promote RPA loading and start HR (Bunting et al., 2010). Although there is no clear evidence could be given for now, the very possible function of BRCA1 and CtlP in this process is to give fine resection of the DSB end to give a hanging over ssDNA end by themselves or by some recruited protein. In BRCA-deficient cells, 53BP1 always binds to the DSB end to inhibit the resection step (Bunting et al., 2010). While, when add 53BP1 loss to the BRCA-deficient cells, there is no more the inhibitory factor 53BP1 block the resection and RPA loading, and the HR function is re-established and even show excess activity than WT cells in some cases (Bunting et al., 2010).

The study on 53BP1 mechanism is very important for the chemotherapy of breast and ovarian cancer. Usually, these cancers are susceptible of mutation in BRCA1 and are treated by PARP inhibitor, which is very toxic for BRCA1-deficient cells (Bunting et al., 2010). However, with the further understanding of 53BP1 function and mechanism, we could develop new therapeutic methods to treat BRCA1 mutation carriers by systemically use of compounds that inhibit 53BP1; this would lead to increased HR activity level in the patient (Cousineau and Belmaaza, 2007). This would be very useful treatment to protect BRCA1 mutation carriers from developing breast cancer, as it is known that heterozygous carrier of cancer mutant gene is very susceptible to gain mutation on the other allele during the tumor progression. Also, this might help to solve the problem of BRCA1 mutant cells developing resistance to PARP inhibitor treatment (Bunting et al., 2010).

Since the outcomes of combination treatment of ATM inhibitor and PARP inhibitors on BRCA1-/- 53BP1-/- cells is consistent with that of 53BP1 inhibitor, it suggested a possible second line of chemotherapy for cancer with resistance to PARP inhibitor by treating with ATM inhibitors (Bunting et al., 2010; Choi et al., 2010). However, there are still many unanswered questions on this regulation mechanism. For example, whether and how 53BP1 restores BRCA2 recruitment and function in BRCA1-/- cells remain unclear. How loss of 53BP1 affects the function of the different distinct BRCA1 associated complexes that are assembled during S-phase and whether this contributes to the “synthetic viable” phenotype is also uncertain (Aly and Ganesan, 2011). We need more investigations on the mechanism of 53BP1 and other ATM related protein to bring these possible chemotherapies into practical use.

Further studies investigated more in the mechanism of 53BP1. It is known that H2AX and 53BP1 co-localize in the nuclear when DSB happens. They illustrated that H2AX also function to inhibit CtlP-mediated DNA end resection (Helmink et al., 2011). Take it into account that the inhibitory effect of 53BP1 on HR DSB repair pathway in cells at post-replicative stages, they proposed the mechanism of H2AX inhibiting DNA end resection by recruiting 53BP1 (Helmink et al., 2011; Bunting et al., 2010). In addition, at the post-replicative stages, H2AX also function to promote DSB repair by HR.

Therefore, H2AX have a more complicated and various roles in regulating the cell cycle (Xie et al., 2007). Meanwhile, another group of researchers investigated the factors that affect 53BP1 function. They found that 53BP1 provide DNA end protection is independent of distance itself, but corresponding to the distance H2AX spreading (Bothmer et al., 2011). Furthermore, other factors, such as chromatin association, oligomerization, and N-terminal ATM phophorylation, are also required for DNA end protection (Bothmer et al., 2011).

In the study of lethality of PARP inhibitor, it was also stated the perspective that disabled NHEJ reduces, rather than exacerbates, the genomic instability and lethality of PARP inhibitor in BRCA1-loss related HR deficient cells. This is contradictory to the original model that HR-deficient cells are dependent on NHEJ pathways to solve the accumulated DNA damage in order to survive under the toxicity of PARP inhibitor (Patel et al., 2011). BRCA1 and 53BP1 could be seen as master regulator of the repair choice that occurs at DNA breaks. That is to say, when 53BP1 is absent, NHEJ is not inhibited and DNA DSB are preferentially repaired by HR; while BRCA1 is absent, DSBs do not undergo NHEJ, resulting in suppression of HR and preferential repair by NHEJ.

When both BRCA1 and 53BP1 are absent, end-resection at DNA DSBs are subsequent HR mediated repair can once again take place (Aly and Ganesan, 2011; Patel et al., 2011). It is shown that PARP inhibitor is only very successful in the early stage of treating BRCA-deficient cancer, and leads to resistance by 33% in BRCA-deficient ovarian cancers (Audeh et al., 2010) and 41% in BRCA-deficient breast cancer (Tutt et al. 2010). However, when comparing to other targeted therapies, these resistance rates are relatively lower (Patel et al., 2011). Therefore, the low NHEJ activity seen in the BRCA-deficient tumors might only affect to reduce the response to PARP inhibitor a little bit.

In conclusion, 53BP1 functions as a regulator in the DSB repair system by blocking the DSB resection process in the HR pathway, and thereby, promoting NHEJ pathway. The relevant mechanism is still under further study and we still have not fully understand what repair factors are regulating end resection and choice of repair pathways in the absence of both 53BP1 and BRCA1 (Aly and Ganesan, 2011). Meanwhile, this is a very useful topic that could lead to practical development in new chemotherapy that could compensate the resistance problem of the first line chemotherapy of PRAP inhibitors.